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Bi W.,Chinese PLA General Hospital | Xiao L.,CAS Institute of Biophysics | Jia Y.,Chinese PLA 306 Hospital | Wu J.,CAS Institute of Biophysics | And 4 more authors.
Journal of Biological Chemistry | Year: 2010

Protein kinases play an important role in the maintenance of homeostasis between cell survival and apoptosis. Deregulation of these kinases leads to various pathological manifestations, such as cancer and neurodegenerative diseases. The MST1 encodes a serine/threonine kinase that is activated upon apoptotic stimulation, which in turn phosphorylates its downstream targets, HistoneH2Band FOXO. However, the upstream regulators of MST1 kinase have been poorly studied. In this study, we report that JNK (c-Jun N-terminal kinase) phosphorylates MST1 at serine 82, which leads to the enhancement of MST1activation.Accordingly,theactivationofMST1phosphorylates FOXO3 at serine 207 and promotes cell death. The inhibition of JNK kinase per se attenuates MST1 activity and nuclear translocation as well as MST1-induced apoptosis. We also find the S82A (serine mutated to alanine) diminishes MST1 activation and its effect on the FOXO transcription activity. Collectively, these findings define the novel feedback regulation of MST1 kinase activation by its putative substrate, JNK, with implication for our understanding of the signaling mechanism during cell death. © 2010 by The American Society for Biochemistry and Molecular Biology, Inc.

Guo H.,Chinese PLA General Hospital | Du G.,Chinese PLA General Hospital | Wang L.,Chinese PLA General Hospital | Wang D.,Chinese PLA General Hospital | And 2 more authors.
Cell Biology International | Year: 2013

The corneal epithelial barrier dysfunction is associated with a number of disorders of the cornea. This study aims to investigate the role of integrin alpha v beta 6 (avb6) in maintaining the corneal epithelial barrier function. In this study, the association of avb6 and endosome/lysosome in the human corneal epithelial cell line, HCE cells, was observed with immunocytochemistry. The corneal epithelial barrier function was assessed with HCE monolayers in Transwells. The results showed that avb6 was observed attaching to endosomes in HCE cells. Knockdown of avb6 resulted in disturbing the fusion of endosome/lysosome. Tumor necrosis factor (TNF)-α could bind avb6 to form a complex that interfered with the fusion of endosome/lysosome in HCE cells. With the HCE monolayer as a corneal epithelial barrier model, the knockdown of avb6 or exposure to TNF-α markedly increased the epithelial barrier permeability to a macomolecular protein, ovalbumin, in Transwell system. After passing through the deficient epithelial barrier, the ovalbumin molecules still conserved the antigenicity. We conclude that the integrin avb6 plays an important role in the fusion of endosome/lysosome in corneal epithelial cells; inhibition of avb6 results in corneal epithelial barrier dysfunction. © 2013 International Federation for Cell Biology.

Li Q.,Capital Medical University | Jie Y.,Capital Medical University | Wang C.,Capital Medical University | Zhang Y.,Capital Medical University | And 2 more authors.
Cell Biochemistry and Function | Year: 2014

Corneal epithelial barrier dysfunction is harmful to corneal health; the pathogenesis is unclear. This study aims to elucidate the mechanism by which tryptase compromises corneal epithelial barrier function. Human corneal epithelial cell line (HCE cells) was cultured into monolayers using as a study platform. Quantitative reverse transcription polymerase chain reaction and Western blotting were employed to detect the expression of matrix metalloprotenases (MMP)9. The endosome/lysosome fusion was observed by confocal microscopy. The corneal epithelial barrier function was assessed in Transwell system. The results showed that HCE cells expressed proteinase-activated receptor (PAR)2. Activation of PAR2 by tryptase induced expression of MMP9 in HCE cells, interfered with the fusion of endosome/lysosome, and compromised the epithelial barrier function, which could be prevented by pretreatment with MMP9 inhibitor. We conclude that tryptase can increase the expression of MMP9 in HCE cells and compromise the epithelial barrier function. © 2013 John Wiley & Sons, Ltd.

Wang L.,Chinese PLA General Hospital | Xu X.,Chinese PLA General Hospital | Huo N.,Chinese PLA General Hospital | Guo H.,Chinese PLA 306 Hospital | And 2 more authors.
Biochemistry and Cell Biology | Year: 2013

Osteocyte generation can be used in bone defect repair; the generation efficiency needs to be further improved. This study aims to evaluate the role of ubiquitin A20 (A20) in facilitating the expression of osteocalcin in adipose-derived stem cells (ADSCs). In this study, adipose tissue was obtained from 10 healthy human subjects; ADSCs were isolated from the adipose samples. The ADSCs were transfected with core binding factor alpha 1 (Cbfa1) and/or insulin-like growth factor-1 receptor (IGF-1R). Expression of osteocalcin, A20 in ADSCs was assessed by quantitative RT-PCR (qRT-PCR) and Western blotting. Apoptosis of ADSCs was analyzed by flow cytometry. The results showed that after the gene transfection and stimulation of insulin, the ADSCs expressed high levels of osteocalcin. However, apoptotic ADSCs were induced by the activation of IGF-1R. Exposure to insulin down-regulated the expression of Bcl-xL and A20, and increased Bax, in ADSCs. The addition of exogenous A20 prevented the ADSC apoptosis. We conclude that activation of IGF-1R can induce apoptosis in ADSCs, which can be prevented by addition of exogenous A20. © 2013 Published by NRC Research Press.

Jinsong C.,Chinese PLA General Hospital | Shanshan Z.,Chinese PLA General Hospital | Zhihong W.,Chinese PLA 306 Hospital | Jing J.,Chinese PLA General Hospital | And 4 more authors.
Experimental and Clinical Cardiology | Year: 2014

Although the incidence of in-stent restenosis (ISR) decreased following the introduction of drug-eluting stents, the absolute number of patients who suffer from ISR has increased. The diagnosis of ISR is dependent on angiography, an expensive and invasive procedure. Therefore, a noninvasive and more affordable method is needed for the detection of ISR. Based on reports that circulating microRNAs are potential biomarkers for various heart diseases, the aim of the current study was to identify microRNAs that could serve as indicators of ISR. Plasma samples from 30 patients with ISR (ISR group) and 30 patients without ISR (NISR group) were collected. Pooled samples of total RNA from each group were screened using a microRNA array, and the results were validated by quantitative reverse transcription-polymerase chain reactions. Seven microRNAs were found to be expressed differentially between the ISR and NISR groups. Further, multivariate analysis revealed that microRNA-21, 100, 143 and 145 were statistically associated with ISR (odds ratio = 0.72, 1.47, 2.41, and 3.24 respectively). We developed a MicroRNA Score based on the cycle threshold values of these 4 microRNAs to predict the occurrence and severity of ISR. The MicroRNA Score was significantly higher in the ISR group than in the NISR group, and was found to discriminate ISR patients from NISR patients, showing a sensitivity of 80% and a specificity of 90%. Moreover, this score had a strong correlation with the severity of restenosis (r value = 0.9290, p = 0.000). These results were validated in a new cohort of 30 patients each in an ISR and NISR group. The combination of microRNA-100, 143, 145, and 21 was able to effectively identify ISR patients and correlated with the severity of restenosis.

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